Astragalus auganus) at Pishin area in Balochistan, Pakistan

Numerous plants of Pakistan are well known for their remedial prospective and are traditionally utilized in herbal medicine system across the country. In the present study, for the first time five of the most prevalent commonly used therapeutic plants (Hertia intermedia, Cardaria chalepense, Scorzonera ammophila, Tamaix karelini and Astragalus. auganus) of Pishin area are studied by atomic absorption spectrometry for their heavy element (Cadmium, Copper, Nickel, Lead and Zinc) concentrations (ppm). Furthermore, some significant features of trace metal toxicity are also deliberated. Though various floras amass heavy metals however our studied plants have low heavy metal concentrations according to the standards of international safety for human consumption. Therefore, all these plants of the area are safe to use for medicinal purpose.


Introduction
Therapeutic plants have always caught the interest of mankind for the treatment of different ailments [1]. These days about 70 -80 % of human population depends upon traditional phytotherapies that are mainly based on folklore system [2]. Particularly the rural populations are still practicing the conventional herbal medicine systems like Ayurvedic, Unani, Chinese and Biomedicine for the treatment of different diseases [3]. Therapeutic plants are also used for primary health care by millions of people in developing and industrialized regions where it is occupying a complementary position for the prevention and treatment of diseases [4]. Therapeutic plants have played significant role in discovering modern medicines with novel organic constituents such as vitamins, glycosides and some essential oils etc. In living organisms such pharmacologically active components are significant for vital physiological processes and curative purposes [4,5]. Accordingly, medical practitioners are also recommending plant medicines, herbal extracts and teas as a complementary treatment for everyday health complications [6]. But the question is that, Are medicinal plants completely safe? While they are often promoted as natural and harmless but herbal preparations are not at all free from adverse properties [7]. Today, it has been a proven fact that persistent ingestion or over dosage of therapeutic herbs causes long-lasting accumulation of several elements in the body which may pose numerous health problems [8]. As remedial herbs preferred for pharmacological actions may be contaminated by heavy metals that cause severe health threats such as symptoms of prolonged toxicity, liver damage, renal failure and certain other unknown allergic or toxic reactions [7,9]. Many herbal medicinal mixtures present dangerous human health threats due to the occurrence of toxic elements like Cd, Hg, Pb, Al and Cr in high concentrations and their oxidation states [10,11]. Though the heavy metals toxicity to human have been identified for a long time however in some areas their exposure is still sustained and even increasing [12,13]. According to the WHO stipulation Pb, Cr, Cd, and other heavy metals must totally be controlled in remedies to assure their safety. Consequently, the discussion on whether herbal medicines should be submitted to appropriate licensing processes integrating quality standards, efficacy and safety is continuing. In such circumstance, the elemental concentrations of therapeutic herbs become very significant, thus their quality control must be properly tested [7,14].
The term heavy metal is substituted by the poorly defined notion of "Trace Elements Metal" (ETM) which include 80 constituents' chemical elements of the crust  [20]. Most heavy metals contaminants do not undergo chemical degradation like organic substances which are degraded to oxides of carbon (IV) microorganisms and their whole concentration after their introduction to soil perseveres for a long time [21]. The biodegradation of organic wastes in soil is severely inhibited by toxic trace metals contamination [22]. This soil contamination may pose threats to humans as well as ecosystem by ingestion or direct contact with the toxic soils, reduced food quality, food chain (soils-plants-animalshuman), phytotoxicity, contaminated drinking water, decrease in land for agricultural production that pose land tenure problems and food insecurity [23,24].
Lead metal is present in group IV and 6 th period. It has atomic number and mass of 82 and 207.2 respectively, while` its melting and boiling points are 327.4 o C and 1725 •C respectively and it has density of 11.4 g/cm 3 . Pb metal is a naturally bluish gray in color and found as a mineral and usually present in combined form with other elements like oxygen (PbCO3) or Sulphur (PbS & PbSO4). It varies in the earth crust from 10 -30 mg /kg [25]. The most stable forms of Pb are Pb II and lead hydroxy complexes. The PbII is the most reactive species of Pb that forms poly nuclear and mononuclear hydroxides and oxides [20]. Pb forms strong oxidant compounds (PbO2) and some basic salts like 2PbCO3, Pb (OH)2 and various organic Pb compounds like tetra methyl or ethyl lead. Lead is positioned 5 th in the industrial production of metals after Fe, Cu, Al and Zn. Hydroxides and oxides of Pb, Ionic lead, oxy-anion complexes of lead and PbII are generally freed into the soil, surface waters and groundwater. Plants generally do not absorb or amass Pb. It is an unessential and toxic element that cause extensive effects of injury to nervous system, brain, kidneys and erythrocytes [26,27]. Zinc is found as a transition metal of group IIB and 4 th period and its atomic number is 30 and atomic mass is 65.4. The melting and billing points of Zn are 419.5 •C and 90 •C respectively, while it has a density of 7.14 gcm -3 . Zn is naturally found in soil (70 mg kg -1 in surface rocks) [28], but its content is increasing due to anthropogenesis like industrial processing, coal and mineral excavation, steel processing and waste burning. Drinking water and many food products have positive Zn concentration that may increase if stored in metallic containers. Ground water and plant may be contaminated by soluble zinc in soils. Plants usually uptake Zn and accumulate higher level than the normal due to its high concentrations in soils [29].
Cadmium is a d' block transition element of group 12 and 5 th period with characteristics: atomic number and weigh of 48 and 112.4 respectively. Its melting and boiling points are 320.9 •C and 765 •C respectively. It has density of 8.65g/cm 3 . Cd is among major heavy metal poisons with mercury and lead. It is found as Cd II ion in its compounds. In plants and animals' cadmium is vital micronutrient. Cd due to chemical similarity can be substituted for Zn which may lead to malfunctioning of metabolic processes [30]. Cadmium is highly bio persistent that remains resident for many years when absorbed by an organism. It reduces the catalytic activity of different enzymes like alcohol or lipoamide dehydrogenase and arylsulfatase, however it enhances the activity of acid dehydratase, deltaamino levulinic, pyruvate decarboxylase and dehydrogenase. Cd is also badly distressing enzymes which reabsorb proteins from kidney tubules resulting proteinuria [31].
Copper is a group IB and 4 th period transition metal with atomic number and weight of 29 and 63.5 respectively. It has density of 8.96 g/cm 3 and melting point and boiling point 1083 •C and 2595 •C respectively [28]. It is the third most used metal in the world [32]. Cu is also a micronutrient that is vital for both animals and plants growth. In humans, Cu aids in the construction of hemoglobin while in plants it is essential in disease resistance, seed making and water regulation. Though Cu is necessary element nonetheless its high concentration is not free of adverse effects of anemia, stomach and intestinal irritation and damage of liver and kidney [33]. Nickel is a d' block transition metal of group 10 and 4 th period with atomic number and weight 28 and 58.69 respectively. It subsists in regions of low pH as Ni II ion, while it precipitates as a stable Ni (OH)2 compound in neutral to slightly alkaline solutions. This precipitate is dissolved in very alkaline solutions to produce nickelite ion, HNiO2 that is solvable in water and in acid conditions to form Ni III [34]. Ni occurs in the environment at low level and is needed in low amounts however if the maximum tolerable quantities are exceeded then Ni can be dangerous that can cause some cancers on different body parts of animals that mainly live near refineries [35]. Hertia intermedia (Boiss) O. Ktze. Also known as Othonnopsis intermedia (Boiss). Hertia intermedia is a small shrub with pretty yellow flowers that belongs to family Asteraceae or Compositae and is a therophytic shrub with microphyllous leaf. It is a Psammophytes that grows at waste water stream margins and sandy gravel slopes [36]. Globally it is found in Buski, Central Asia and Vern. The genus Hertia, has 12 species found all over southwest Asia and north and south Africa [37]. In Pakistan it is found in some areas of province Baluchistan where it is common in Quetta, Koeie, Chaman, Pishin (Kanozai area), and Wazir. It is also found in Kurram and regions below Parachinar. it is locally named as Gaungha. This plant blooms up in is April. The leaves of Hertia intermedia are of medicinal importance and are traditionally used for boils, pimples and headache treatments. The general image of Hertia intermedia is given bellow in figure 1. Cardaria chalepense (Linnues) belongs to family Brassicaceae and it is a therophyte ruderal herb having microphyll leaf size. The Brassicaceae is a family of 350 genera and about 3000 species that are mainly found in cooler and temperate climates [38,39]. Globally it is found in central Asia and in Pakistan Cardaria chalepense is found in Pishin, Sorab, Kalat and Quetta. About 92 genera and 250 species are recorded from our area that contain 5 genera and 14 species identified from cultivation [40]. Locally it is named as Bashki and the blooming season is April-July. Cardaria chalepense (Bashki) is well known for its peppery tasted oil producing seeds and use as a green fodder and vegetable. This     Table 1. Samples collection of the selected plants were carried out from three different villages named as Tora Ghundai, Batazai and lamarran that lie adjacent to the main city of Pishin district. Random samples from each site were collected in sterile plastic bags and were conveyed to our Laboratory of chemistry department (UOB) for further examinations. These plants were identified, and their nomenclature was ascertained on basis of the flora of Pakistan [51]. Sample preparation All samples of plants were thoroughly washed by deionized water. Then these washed samples were oven dried for 2 days at 80°C and then grinded the dry matter of the whole plant parts into powder form. Samples were precisely weighed (0.5g each) and placed in a 100mL PTFE beaker. The samples were then subjected to acid digestion method that is explained below to determine the concentrations (ppm) of Cd, Cu, Ni, Pb and Zn in samples. Then about 3ml of sample solutions of each plant was used to calculate the unknown concentration of the five selected heavy metals. Only Cd, Cu, Ni, Pb and Zn form all heavy metals were selected on basis of lab facilities and other heavy metals were skipped.

Results and discussion
All over the world the concentrations of both essential and non-essential trace heavy elements in therapeutic herbs beyond the approved limit is a great matter of concern for public safety. In Pakistan this problem is even more serious and alarming due to uncontrolled and no proper regulation of quality assurance parameters of plants which form the raw materials for the finished therapeutic products.

Cadmium
In the studied plants, the high concentration of Cd in ppm was observed to be 0.0338 in Scorzonera ammophila followed by 0.012 in Cardaria chalepense, 0.012 in Tamarix karelini, 0.008 in Hertia intermedia and 0.005 in Astragalus auganus (Table 2 & Figure 6). The World Health Organization/ Food / Agriculture Organization [53] has set 0.21 ppm as permissible limit for Cd in edible plants. Though

Conclusion
The five selected therapeutic plants have been suggested as remedies for numerous disorders in the conventional medicine system. There has been a great progress about scientific evaluation of therapeutic herbs in the field of phytotherapy throughout the world. The practical impact of the changing conditions may be witnessed in the WHO monographs, national pharmaceutical products and herbs dealing industries. The trace heavy metals contents of studied therapeutic plants were determined to be well lower than the critical limits.